Phosphorylation of protein kinase C delta on distinct tyrosine residues induces sustained activation of Erk1/2 via down-regulation of MKP-1 - Role in the apoptotic effect of etoposide

The mechanism underlying the important role of protein kinase C delta (PKC delta) in the apoptotic effect of etoposide in glioma cells is incompletely understood. Here, we examined the role of PKC delta in the activation of Erk1/2 by etoposide. We found that etoposide induced persistent activation of Erk1/2 and nuclear translocation of phospho-Erk1/2. MEK1 inhibitors decreased the apoptotic effect of etoposide, whereas inhibitors of p38 and JNK did not. The activation of Erk1/2 by etoposide was downstream of PKC delta since the phosphorylation of Erk1/2 was inhibited by a PKC delta-KD mutant and PKC delta small interfering RNA. We recently reported that phosphorylation of PKC delta on tyrosines 64 and 187 was essential for the apoptotic effect of etoposide. Using PKC delta tyrosine mutants, we found that the phosphorylation of PKC delta on these tyrosine residues, but not on tyrosine 155, was also essential for the activation of Erk1/2 by etoposide. In contrast, nuclear translocation of PKC delta was independent of its tyrosine phosphorylation and not necessary for the phosphorylation of Erk1/2. Etoposide induced down-regulation of kinase phosphatase-1 (MKP-1), which correlated with persistent phosphorylation of Erk1/2 and was dependent on the tyrosine phosphorylation of PKC delta. Moreover, silencing of MKP-1 increased the phosphorylation of Erk1/2 and the apoptotic effect of etoposide. Etoposide induced polyubiquitylation and degradation of MKP-1 that was dependent on PKC delta and on its tyrosine phosphorylation. These results indicate that distinct phosphorylation of PKC delta on tyrosines 64 and 187 specifically activates the Erk1/2 pathway by the down-regulation of MKP-1, resulting in the persistent phosphorylation of Erk1/2 and cell apoptosis.